Ingestion of microorganisms by neutrophils triggers a complex cellular response that includes activation of a superoxide-generating oxidase, an acidification process, and profound selective alterations of membrane permeability. These responses are activated within 30-60 seconds of exposure to the initiating soluble or particulate stimulus. Preliminary experiments in our laboratory strongly suggest that multistep transducing sequences are interposed between the initial membrane perturbation caused by contact with the stimulus and the subsequent responses of the neutrophil. Since such neutrophil responses play important roles in states of inflammation and infection, we have designed this research proposal to characterize the transducing sequences and identify their components. As a necessary component of this endeavor, we will also define more completely the altered permeability characteristics and acidification mechanisms of activated neutrophils. To accomplish these objectives, suspensions and monolayers of purified human neutrophils will be examined under basal and stimulated conditions. Phagocytic stimuli will include heat-killed microorganisms and polystyrene latex particles. Soluble stimuli will include digitonin and the ionophore A 23187. Oxidase activation will be assessed by measuring O2 consumption and O2 (superoxide) production. Membrane permeability will be assessed by measuring the influx and efflux of physiologically significant cations (K ion, Rb ion, Na ion, H ion, and Ca 2 ion) and selected amino acids and hexoses. We will use various inhibitors to "dissect" the transducing sequence. Prominent among these inhibitors will be agents known to affect microtubules, microfilaments, phospholipases, membrane ATPases and calcium transport. Standard metabolic inhibitors (e.g. cyanide, 2-deoxyglucose, sulfhydryl reagents, etc.) will also be used. As dictated by laboratory findings with normal neutrophils, selected studies will be performed on neutrophils from subjects with chronic granulomatous disease, hereditary myeloperoxidase-deficiency, and other states of leukocyte dysfunction.